• Abstract and Figures
• Public Full-text

Pancreatic carboxyl ester lipase: a circulating enzyme that modifies normal and oxidized lipoproteins in vitro. R Shamir, … , D W Morel, E A Fisher J Clin Invest. 1996;97(7):1696-1704. https://doi.org/10.1172/JCI118596. Research Article Pancreatic carboxyl ester lipase (CEL) hydrolyzes cholesteryl esters (CE), triglycerides (TG), and lysophospholipids, with CE and TG hydrolysis stimulated by cholate. Originally thought to be confined to the gastrointestinal system, CEL has been reported in the plasma of humans and other mammals, implying its potential in vivo to modify lipids associated with LDL, HDL (CE, TG), and oxidized LDL (lysophosphatidylcholine, lysoPC). We measured the concentration of CEL in human plasma as 1.2+/-0.5 ng/ml (in the range reported for lipoprotein lipase). Human LDL and HDL3 reconstituted with radiolabeled lipids were incubated with purified porcine CEL without or with cholate (10 or 100 microM, concentrations achievable in systemic or portal plasma, respectively). Using a saturating concentration of lipoprotein-associated CE (4 microM), with increasing cholate concentration there was an increase in the hydrolysis of LDL- and HDL3-CE; at 100 microM cholate, the present hydrolysis per hour was 32+/-2 and 1.6+/-0.1, respectively, indicating that CEL interaction varied with lipoprotein class. HDL3-TG hydrolysis was also observed, but was only approximately 5-10% of that for HDL3-CE at either 10 or 100 microM cholate. Oxidized LDL (OxLDL) is enriched with lysoPC, a proatherogenic compound. After a 4-h incubation with CEL, the lysoPC content of OxLDL was depleted 57%. Colocalization of CEL in the vicinity of OxLDL formation was supported by demonstrating in human aortic homogenate a cholate-stimulated […] Find the latest version: https://jci.me/118596/pdf Pancreatic Carboxyl Ester Lipase: A Circulating Enzyme That Modifies Normal and Oxidized Lipoproteins In Vitro Raanan Shamir,*‡ William J. Johnson,* Kelly Morlock-Fitzpatrick,* Reza Zolfaghari,* Ling Li,§ Eric Mas,ʈ Dominique Lombardo,ʈ Diane W. Morel,* and Edward A. Fisher*§ *Department of Biochemistry, Medical College of Pennsylvania and Hahnemann University, Philadelphia, Pennsylvania 19129; ‡Division of Gastroenterology and Nutrition, The Children’s Hospital of Philadelphia, Philadelphia, Pennsylvania 19104; §Cardiovascular Institute, Mount Sinai School of Medicine, New York 10029; and ʈINSERM U260, Faculté de Médicine, 13385 Marseille, France Abstract Introduction Pancreatic carboxyl ester lipase (CEL) hydrolyzes choles- Modification by lipases, such as hepatic lipase (1), lipoprotein teryl esters (CE), triglycerides (TG), and lysophospholipids, lipase (2), and phospholipase A2 (3), alters the composition, with CE and TG hydrolysis stimulated by cholate. Origi- structure, and function of lipoproteins. Neglected in this re- nally thought to be confined to the gastrointestinal system, gard is the pancreatic carboxyl ester lipase (CEL,1 sometimes CEL has been reported in the plasma of humans and other referred to as bile salt–dependent cholesteryl ester hydrolase), mammals, implying its potential in vivo to modify lipids as- whose primary function is thought to be hydrolysis of dietary sociated with LDL, HDL (CE, TG), and oxidized LDL (lyso- lipid esters in the intestinal lumen (for a recent review, see ref- phosphatidylcholine, lysoPC). We measured the concentra- erence 4). tion of CEL in human plasma as 1.2Ϯ0.5 ng/ml (in the One reason for this neglect may be the lack of appreciation range reported for lipoprotein lipase). Human LDL and that CEL activity is expressed outside of the pancreas in a HDL3 reconstituted with radiolabeled lipids were incubated number of mammalian species. For example, there are reports with purified porcine CEL without or with cholate (10 or of CEL activity and/or mRNA in rat liver (e.g., references 100 ␮M, concentrations achievable in systemic or portal 5–7), human breast (8), human hepatocarcinoma HepG2 cells plasma, respectively). Using a saturating concentration of (9), human placenta (10), rat (11) and rabbit heart (12), and rat lipoprotein-associated CE (4 ␮M), with increasing cholate and rabbit aorta (13, 14). Particularly intriguing is the evidence concentration there was an increase in the hydrolysis of that CEL is found in the blood plasma of a number of mam- LDL- and HDL3-CE; at 100 ␮M cholate, the percent hydro- mals, including humans (15), rats (11), dogs, and goats (16). lysis per hour was 32Ϯ2 and 1.6Ϯ0.1, respectively, indicat- Implied, then, is that lipoproteins in the circulation or intersti- ing that CEL interaction varied with lipoprotein class. tium could be potential targets for CEL and also that circulat- ف HDL3-TG hydrolysis was also observed, but was only 5– ing CEL, which has a heparin binding site (17), may become 10% of that for HDL3-CE at either 10 or 100 ␮M cholate. associated with a variety of tissues, independent of their capac- Oxidized LDL (OxLDL) is enriched with lysoPC, a pro- ity to synthesize the enzyme, by associating with cell-surface atherogenic compound. After a 4-h incubation with CEL, proteoglycans. The consequences of CEL-mediated lipopro- the lysoPC content of OxLDL was depleted 57%. Colocal- tein modification are suggested by recent reports that (a) the ization of CEL in the vicinity of OxLDL formation was sup- treatment in vitro of LDL by nonmammalian cholesteryl es- ported by demonstrating in human aortic homogenate a terases depletes LDL of core cholesteryl esters (18) and leads cholate-stimulated cholesteryl ester hydrolytic activity in- to increased net transfer of LDL-cholesterol to cells (19), and hibited by anti–human CEL IgG. We conclude that CEL (b) CEL secreted by HepG2 cells increases the cellular uptake has the capability to modify normal human LDL and HDL of cholesteryl esters from HDL (20). composition and structure and to reduce the atherogenicity In addition to its activity against the dietary lipids choles- of OxLDL by decreasing its lysoPC content. (J. Clin. Invest. teryl ester (CE), triglyceride (TG), and retinyl ester, CEL has 1996. 97:1696–1704.) Key words: low density lipoprotein • significant activity against lyosophospholipids and, in fact, was high density lipoprotein • oxidized low density lipoprotein • originally cloned as the pancreatic lysophospholipase (21). aorta • lysophosphatidyl choline Thus, it is plausible that CEL may modify not only the core CE or TG of a normal lipoprotein, but also the lysophosphatidyl- choline (lysoPC) of oxidized LDL (OxLDL). Given the major roles attributed to lysoPC in the promotion of atherosclerotic disease (such as enhanced monocyte chemotaxis [22], mito- genic stimulation of macrophages [23], and inhibition of endo- Address correspondence to Dr. Edward A. Fisher, Mount Sinai Car- thelium-dependent arterial relaxation [24]) and the reports diovascular Institute, The Rockefeller University, Box 4, 1230 York that CEL activity was found in rat and rabbit aortae (13, 14), Ave., New York, NY 10021. Phone: 212-327-7452; FAX: 212-327- CEL could serve as a protective factor in vascular tissue 7454; E-mail: [email protected] Received for publication 21 September 1995 and accepted in re- vised form 16 January 1996. 1. Abbreviations used in this paper: CE, cholesteryl ester; CEL, car- J. Clin. Invest. boxyl ester lipase; HDL3, HDL, subclass 3; LpL, lipoprotein lipase; © The American Society for Clinical Investigation, Inc. lysoPC, lysophosphatidylcholine; OxLDL, oxidized LDL; PC, phos- 0021-9738/96/03/1696/09 $2.00 phatidylcholine; TBARS, thiobarbituric acid–reactive substances; Volume 97, Number 7, April 1996, 1696–1704 TG, triglyceride or triacylglycerol. Lipoprotein Modification by Pancreatic Carboxyl Ester Lipase 1696 against the adverse effects of OxLDL. Supporting this hypoth- phoresis of native and reconstituted lipoproteins and gel staining by esis are the recent reports that the treatment of OxLDL with a Sudan black were performed using Beckman Paragon Lipo Gel mate- bacterial lysophospholipase reduced the antagonism by rials. lysoPC of the endothelial-dependent relaxation response to Preparation and characterization of oxidized human LDL acetylcholine of aortic rings (24, 25). Since oxidation of LDL is thought to occur not in plasma but within arterial tissue (26), Human LDL, isolated as above, was oxidized by a 7-h incubation with Cu2+ (5 ␮M above EDTA concentration [37]). Butylated hy- aortic CEL activity would be in the appropriate location to droxytoluene (20 ␮M) was then added to stop any further oxidation. serve as a protective factor. The cholesterol content of LDL and OxLDL was determined enzy- In the studies summarized in this report, we have demon- matically by the Sigma cholesterol diagnostic kit No. 352. Protein strated: (a) the ability of CEL to hydrolyze the core lipids of content was assayed using a modification of the Lowry technique human LDL and HDL using bile salt concentrations that can (35). The cholesterol contents of LDL and OxLDL were 1.4 mg and occur outside of the intestinal lumen in healthy individuals; (b) 1.2 mg/mg protein, respectively. The protein concentrations of LDL the ability of CEL to reduce the lysoPC content of human Ox- and OxLDL were 1.4 and 1.3 mg/ml, respectively. The extent of lipid LDL; (c) the presence of CEL in human plasma; and (d) the peroxidation was estimated as thiobarbituric acid–reactive substances presence of an activity in human aortic homogenate with enzy- (TBARS) as described previously (38). Briefly, LDL or OxLDL (50 ␮ matic and immunological properties indistinguishable from g protein of either) and various amounts of standard (1,1,3,3-tet- ramethoxypropane) were mixed with 1 ml of 25% trichloroacetic acid CEL. The results strongly suggest that CEL plays a wider role and 1 ml of 1% 2-thiobarbituric acid containing 5 ␮M CuSO4. The in lipid and lipoprotein metabolism than previously appreci- mixture (final volume of 0.4 ml) was incubated at 95ЊC for 45 min and ated both in terms of enzymatic targets and tissue distribution. centrifuged at 1,500 rpm at 10ЊC for 30 min to remove any insoluble material.

Load more

  10

Copy Link